Organophosphate and organophosphonate esters are among the most toxic compounds that have ever been synthesized. These compounds are toxic because of their inherent ability to interfere with nerve transmission through the rapid and irreversible inactivation of the enzyme acetyl cholinesterase. The toxic properties of the organophosphates have been exploited as agricultural insecticides and as chemical weapons. A bacterial phosphotriesterase enzyme has been identified that can function as a catalyst for the recognition, hydrolysis, and detoxification of a broad spectrum of organophosphate nerve agents. The primary objective of this application is to design and characterize modified forms of the bacterial phosphotriesterase that are optimized for the selective recognition and destruction of those organophosphates that pose the most serious threats to human health. Rational and combinatorial libraries of mutant enzymes will be constructed and those variants with enhanced catalytic proficiency will be identified through novel screening and selection procedures. The progressive changes in the amino acid sequence of the bacterial phosphotriesterase will be directly correlated with the enhancements in the catalytic constants and the modifications within the active site as determined by X-ray diffraction methods. PUBLIC HEALTH RELEVANCE: The toxic properties of the organophosphate nerve agents represent a serious threat to the health and well being of civilized societies. The primary objective of this project is to create novel catalytic enzymes that are optimized in their ability to detect, destroy and detoxify a broad spectrum of organophosphate nerve agents.